High-Throughput Profiling of Anti-Glycan Humoral Responses to SIV Vaccination and Challenge

Abstract

Recent progress toward an HIV vaccine highlights both the potential of vaccines to end the AIDS pandemic and the need to boost efficacy by incorporating additional vaccine strategies. Although many aspects of the immune response can contribute to vaccine efficacy, the key factors have not been defined fully yet. A particular area that may yield new insights is anti-glycan immune responses, such as those against the glycan shield that HIV uses to evade the immune system. In this study, we used glycan microarray technology to evaluate anti-glycan antibody responses induced by SIV vaccination and infection in a non-human primate model of HIV infection. This comprehensive profiling of circulating anti-glycan antibodies found changes in anti-glycan antibody levels after both vaccination with the Ad5hr-SIV vaccine and SIV infection. Notably, SIV infection produced generalized declines in anti-glycan IgM antibodies in a number of animals. Additionally, some infected animals generated antibodies to the Tn antigen, which is a cryptic tumor-associated antigen exposed by premature termination of O-linked glycans; however, the Ad5hr-SIV vaccine did not induce anti-Tn IgG antibodies. Overall, this study demonstrates the potential contributions that glycan microarrays can make for HIV vaccine development.

Serum glycan-binding IgG antibodies in HIV-1 infection and during the development of broadly neutralizing responses

“…All human serum samples were heat-inactivated for 30 minutes in a 56°C water bath prior to binding on the glycan arrays. Similar experiments using samples from macaques showed no inhibitory effect on antibody function or anti-glycan antibody profiles [32, 33] . All time-points from the same individual were run on the same day to minimise intra-individual variability and samples from individuals with and without bNAbs were intermixed to minimise experimental biases.…”

“…Increased binding to Tn-peptides has also been observed following SIV vaccination and SIV infection using this same array [26] . Tn-peptides are precursors of O -linked glycans and in normal mammalian cells the Tn-portion of the O -linked glycans are masked during the glycosylation pathway.…”

“…Tn-peptides are precursors of O -linked glycans and in normal mammalian cells the Tn-portion of the O -linked glycans are masked during the glycosylation pathway. However, in HIV infection the normal glycosylation pathways are altered revealing the Tn-portion of these O -linked glycans on the glycan shield potentially rendering them more immunogenic [26] . Interestingly, anti-Tn-peptide antibodies have been associated with the neutralization of HIV-1 by preventing fusion of the virus into infected cells [46, 47] .…”

“…Included in the array were ten controls (BSA, HSA, linkers and fluorochromes) that were used to assess technical reproducibility and were excluded from downstream analysis. The arrays were generated as previously described [32] with the exception of the addition of a washable fluorescent dye, DyLight™649 (0.7 μg/mL, ThermoScientific) to the print buffer as an indicator of successful liquid deposition and spot morphology. Triton X-100 concentration was reduced to 0.0001% instead of 0.006% to produce smaller spots allowing additional glycans to be spotted onto the plate.…”

“…In addition, the anti-glycan antibody responses in sera of non-human primates after Ad5hr-SIV vaccination and SIV infection [26] and HIV vaccine responses in rabbits have been assayed on arrays [20, 27, 28] . Here we studied serum anti-glycan IgG antibody profiles of HIV negative and HIV positive individuals, including those who develop bNAbs, over three years of infection.…”